DESCRIPTION Recent, advances in nuclear transfer in cattle indicate that it is possible to genetically modify cultured fetal fibroblasts and use them as nuclear donor for the generation of a transgenic offspring. The overall goal of my laboratory is to generate transgenic cattle and pigs with precise genetic modifications and to use the generated animals in biomedical research. As such the aims of the proposed research concentrate on the generation of transgenic bioreactors by combining genetic manipulation of cultured cells and nuclear transfer/cloning of fetal fibroblasts. We are proposing to express the human myelin basic protein (MBP) in the milk as a source of antigen for induction of oral tolerance to MBP and the amelioration of pathologies associated with muscular dystrophy. The project will be divided into two aims: 1) Generation of transgenic fetal bovine fibroblasts with a construct carrying the human myelin basic protein under the control of the whey acidic protein promoter. This initial aim will allow the rapid reestablishment of procedures for culture and manipulation of bovine fetal fibroblasts and the initiation of nuclear transfer with transgenic cells. 2) The second aim will entail the introduction of the myelin basic protein gene into the casein locus by homologous recombination. The purpose of this aim to use the endogenous regulatory sequences of the casein gene to express the MBP gene. Both the MBP cassette and the casein genomic sequence has been isolated and will be available for the generation of the targeting constructs. This aim will allow the development of technology required for targeting specific genes in cultural fetal fibroblasts.